Kit employs the proven methodology for RNA sample preparation and labeling based on the original T7 in vitro transcription (IVT) amplification technology, known as the Eberwine or reverse transcription-IVT (RT-IVT) method. The RT-IVT method is considered the gold standard for sample preparation in microarray-based expression profiling. It is well documented in the current scientific literature and was experimentally validated using TaqMan RT-PCR. Builds upon the linear RNA amplification method and reagents developed for MessageAmp II Kits with enhancements that create a simplified workflow, improved performance, and more flexible RNA input requirements.

Using the MessageAmp Premier RNA Amplification KitBased on this proven enzymatic amplification and labeling strategy, the MessageAmp Premier procedure starts with a simple cDNA synthesis reaction using ArrayScript reverse transcriptase. No cleanup step is required after the RT; cDNA is used directly in a high yield IVT reaction using MEGAscript technology. The IVT is configured to incorporate the modified nucleotide, biotin-UTP, into the aRNA synthesized. (In this protocol, the amplified RNA is referred to as aRNA; in the literature, it is also commonly called cRNA.) Once purified, the aRNA is suitable for use on microarray gene expression analysis platforms designed for biotin-modified antisense RNA samples.

Very flexible RNA input requirements: 20-500ng total RNA, depending on the tissue or cell type

Streamlined workflow: depending on the RNA input amounts, only a single day of processing time is required to complete a microarray experiment

Simplified protocol and streamlined reagents

Complete kit: everything needed to prepare biotin-modified aRNA is included

Rigorous quality control that includes GeneChip analysis

T7 MEGAscript IVT delivers up to 50,000-fold amplification

Single-tube format

Easy-to-use, high-recovery, magnetic-bead aRNA purification

Lowest RNA input requirements for a single round of amplification:

20ng total RNA yields enough biotin-modified aRNA for most microarray formats, with results equivalent to those from samples prepared from significantly more starting material using other RT-IVT kits

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